Nutrient homeostasis is maintained through a complex regulatory network formed by signaling and transcriptional components that control metabolic genes. The liver is one of the key tissues that depending of the physio/pathological conditions buffers whole body nutrient homeostasis. Insulin is one of the most powerful hormones that affect nutrient regulation and clinically, insulin resistance is a hallmark of the metabolic syndrome including type 2 diabetes. As a consequence of insulin resistance one of the metabolic processes that contribute to maintain the diabetic state is hepatic glucose production that is controlled, at least in part, at the transcriptional level. The PI3K/Akt pathway is one of the main effectors of insulin metabolic action. Akt controls expression of metabolic genes through direct phosphorylation and negative regulation of the forkhead transcription factor FoxO1 and coactivators such as PGC-11 and CRCT2, key components of the transcriptional gluconeogenic program. Although Akt can directly mediate this action, there are conditions such as late refeeding or diabetic states where active Akt does not entirely correlate with suppression hepatic glucose production. This indicates that additional key regulatory components, likely kinases, could mediate this repression. Along these lines, we have recently identified Cdc2-like kinase 2 (Clk2) as a novel component downstream of insulin/Akt and functions as part of the hepatic feeding response. Notably, Clk2 controls expression of gluconeogenic genes, hepatic glucose output and blood glucose levels. Moreover, obese/diabetic db/db mice have lower amounts of Clk2 protein and restoration of the levels corrects hyperglycemia. This result suggests that Clk2 might be dysregulated in conditions of obesity/diabetes and contributes to the clinical manifestations. Based on these findings, the major goal of this proposal is to identify the molecular mechanisms by which insulin controls Clk2 kinase activity and to test Clk2 metabolic functionality in-vitro and in in-vivo mouse models. We have three Specific Aims: Aim 1 is to perform molecular and functional analysis of how insulin controls Clk2 kinase activity focusing on regulation of Clk2 protein degradation through ubiquitin ligases. Aim 2 is devoted to carry out molecular and functional analysis of Clk2- mediated suppression of hepatic gluconeogenesis. We focus on the Clk2-induced phosphorylation and repression of PGC-11. Aim 3 will determine the effects of Clk2 on hepatic glucose and lipid metabolism in mice through loss-of-function of hepatic Clk2 in fasting/feeding cycles and genetic and diet-induced obesity. The outcomes of these studies will provide the identification of the molecular mechanisms by which insulin controls glucose and lipid metabolic effects through Clk2 kinase. Since insulin resistance and as a consequence increased and uncontrolled hepatic glucose output is a major defect that occurs in type 2 diabetes, our investigation on the regulation of Clk2 and PGC-11 might translate into potential therapies to treat this disease. PUBLIC HEALTH RELEVANCE: Insulin resistance and increased hepatic glucose production are hallmarks of metabolic diseases such as obesity and type 2 diabetes, thus studies in this grant proposal focusing on key regulators of these processes including a novel kinase Clk2 and PGC-11 might translate into potential therapies.